Effect of abscisic acid, osmolarity and partial desiccation on the development of recalcitrant mango somatic embryos

Inhibition of mango somatic embryo growth was inducedin vitro by treatments for 4 or more weeks with abscisic acid (0–100 M ABA) with and without high osmolarity provided by mannitol (0–10%). High osmolarity and ABA significantly affected somatic embryo length, precocious germination and the production of good quality secondary somatic embryos. High osmolarity also affected root elongation. Abscisic acid was more effective in suppressing growth and development of 0.5 cm-length somatic embryos than smaller somatic embryos. Development beyond the heart stage was significantly inhibited by both ABA and mannitol treatments. The recovery of good quality somatic embryos was enhanced by high levels of ABA (100 M) with and without mannitol (0–5%). Somatic embryos that had been pulsed with ABA were partially desiccated at different relative humidities. Weight loss was affected only by relative humidity; and ABA did not enhance desiccation tolerance.Abbreviations ABA Abscisic acid - 2,4-D 2,4-Dichlorophenoxyacetic acid - MM1 Mango maturation medium - RH Relative humidity     

Control of hyperhydricity of mango somatic embryos

Hyperhydricity of immature somatic embryos has been a limiting factor for the development of highly embryogenic suspension cultures of many important mango cultivars. Reversion of hyperhydricity was achieved in two ways: 1) heart-stage somatic embryos (2–3 mm length) were partially dehydrated under controlled conditions at high relative humidity (RH) for 24–48 h and 2) the gelling agent (Gel-Gro) concentration of the plant growth medium was increased from 2.0 to 6.0 g l^-1. Partially dehydrated immature somatic embryos were normal in appearance. Somatic embryos that were partially dehydrated germinated precociously when cultured on maturation medium. Although abscisic acid (ABA) did not reverse hyperhydricity of primary somatic embryos, ABA did stimulate the reversal of this abnormal pattern of development among secondary embryos. ABA (500 M) inhibited precocious germination and permitted somatic embryo maturation. Partially dehydrated, immature somatic embryos (4–7 mm long) remained viable for up to 32 days in the absence of maturation medium under high RH.Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid - ABA abscisic acid - BA 6-benzyladenine - RH relative humidity     

Effect of exogenous ABA on embryo maturation and quantification of endogenous levels of ABA and IAA in <Emphasis Type="Italic">Quercus suber</Emphasis> somatic embryos

Knowledge of the relationship between indole-3-acetic acid (IAA) and abscisic acid (ABA) is relevant to control the development and the maturation of cork oak (Quercus suber L.) somatic embryos. The addition of 1 M ABA to the culture medium significantly promoted somatic embryo maturation and increased both fresh and dry matter without affecting the relative water content. This effect was parallel to the pattern of variation observed in the endogenous ABA level, which increased from the immature to the mature stage. Endogenous ABA content during the occurrence of secondary embryogenesis was similar to that of the immature stage, showing that embryos with lower ABA levels produced secondary embryos. In contrast, IAA showed the highest concentration during early embryo development and decreased afterwards. Only in somatic embryos subjected to 1-week desiccation followed by stratification at 4 °C for 2 weeks, was a moderate increment of endogenous IAA content observed. IAA and ABA showed opposite levels during the development and maturation of cork oak somatic embryos and characterised specific stages of the embryonic development.     

Manipulating the desiccation tolerance and vigor of dry somatic embryos of Medicago sativa L. with sucrose,heat shock and abscisic acid

Summary The effects of sucrose concentration in the maturation medium in combination with a heat shock treatment at 36°C were investigated in an attempt to improve the vigor of seedlings grown from dry somatic embryos of alfalfa (Medicago sativa L.). Callus was formed from petiole expiants and dispersed in liquid suspension medium in the presence of 5 M 2,4-D. The cell suspension was sieved to synchronize embryo development. The 200 – 500 m fraction was plated on embryo development medium without 2,4-D, grown for 14 days, and transferred to maturation medium. With 3% sucrose in the maturation medium, the somatic embryos germinated precociously and were unable to survive desiccation. At higher sucrose concentrations, germination was delayed and the embryos continued to accumulate dry matter. After 13 days on 6% sucrose medium (27 days after sieving), the somatic embryos were tolerant of drying to 12% moisture without exposure to exogenous ABA. Heat shock, which presumably stimulates endogenous ABA synthesis, improved the desiccation tolerance of somatic embryos if applied prior to day 27 after sieving, but its effects were minimal after day 27. High sucrose concentrations up to 9% in the maturation medium were optimal during the first 8 days on maturation medium (days 14 to 22 after sieving), but a lower concentration (6%) was optimal during the later stages of embryo maturation (days 22 to 30 after sieving). The inclusion of 10^–5 M ABA in the maturation medium with 6% sucrose further improved embryo quality if applied approximately 20 days after sieving.     

Manipulation of conditions for the culture of somatic embryos of white spruce for improved triacylglycerol biosynthesis and desiccation tolerance

In order to enhance post-germinative vigour, somatic embryos of Picea glauca (Moench) Voss. were matured under in-vitro conditions that stimulated triacylglycerol (TAG) biosynthesis. In P. glauca seeds over 90% of the TAG was stored within the megagametophyte, and isolated zygotic embryos contained twice the amount of TAG of somatic embryos cultured for four weeks on basal medium containing 16 M abscisic acid (ABA). Polyethylene glycol-4000 (PEG) as a non-permeating osmoticum with ABA promoted TAG biosynthesis by somatic embryos and sustained maturation throughout an eight-week culture period. Treatments that promoted TAG biosynthesis also prevented precocious germination and promoted desiccation tolerance. Thus, the optimal culture conditions for maturation, desiccation survival, and plantlet regeneration were 16–24 M ABA and 7.5% PEG for eight weeks, followed by desiccation. Under these conditions the levels of TAG per somatic embryo were raised ninefold to about five times the zygotic-embryo level, and the TAG fatty-acid composition became similar to that of zygotic embryos. A study of sectioned material, using light and transmission electron microscopy, showed that the structure and distribution of lipid bodies within these somatic embryos and the degree of embryo development were similar to mature zygotic embryos. Up to 81% of the desiccated somatic embryos regenerated to plantlets during which time the TAG was utilised in a manner similar to zygotic seedlings.Abbreviations ABA abscisic acid - PEG polyethylene glycol - TAG triacylglycerol - TL total lipid - TEM transmission electron microscopy Plant Research Centre contribution No. 1383We are grateful to Dawn Moore and Ken Stanley for technical assistance, and thank Pat Rennie for the electron microscopy. We acknowledge financial support through an NSERC/Forestry Canada/Weyerhaeuser Canada Ltd (Prince Albert, Sask.) research partnership programme.     

In vitro somatic embryogenesis from cell suspension cultures of cowpea [Vigna unguiculata (L.) Walp]

We report, an efficient protocol for plantlet regeneration from the cell suspension cultures of cowpea through somatic embryogenesis. Primary leaf-derived, embryogenic calli initiated in MMS [MS salts (Murashige and Skoog 1962) with B5 (Gamborg et al. 1968) vitamins] medium containing 2,4-Dichlorophenoxyacetic acid (2,4-D), casein hydrolysate (CH), and l-Glutamic acid-5-amide (Gln). Fast-growing embryogenic cell suspensions were established in 0.5 mg l^–1 2,4-D, which resulted in the highest recovery of early stages of somatic embryos in liquid MMS medium. Embryo development was asynchronous and strongly influenced by the 2,4-D concentration. Mature monocotyledonary-stage somatic embryos were induced in liquid B5 medium containing 0.1 mg l^–1 2,4-D, 20 mg l^–1 l-Proline (Pro), 5 M Abscisic acid (ABA), and 2% mannitol. B5 medium was found superior for the maturation of somatic embryos compared to MS and MMS media. The importance of duration (5 d) for effective maturation of somatic embryos is demonstrated. A reduction in the 2,4-D level in suspensions increased the somatic embryo induction and maturation with decreased abnormalities. Sucrose was found to be the best carbon source for callus induction while mannitol for embryo maturation and maltose for embryo germination. Extension of hypocotyls and complete development of plantlet was achieved in half-strength B5 medium supplemented with 3% maltose, 2500 mg l^–1 potassium nitrate, and 0.05 mg l^–1 thidiazuron (TDZ) with 32% regeneration frequency. Field-established plants were morphologically normal and fertile. This regeneration protocol assures a high frequency of embryo induction, maturation, and plantlet conversion.     

The effects of abscisic acid on the maturation ofBrassica napus somatic embryos

Summary A comparison of embryos, cultured for increasing periods of time with and without abscisic acid (ABA), was undertaken to investigate, at the ultrastructural level, the influence of this growth regulator on the maturation of rapeseed (Brassica napus) somatic embryos. In the absence of ABA, the embryos germinated precociously while lipid bodies (LB), which were not numerous, soon degraded, as revealed by a depletion process associated with the appearance of morphologically mature glyoxysomes and an increase in the number of mitochondria. Moreover, a lack of protein bodies indicated that storage protein accumulation was not initiated under these conditions. On the contrary, the addition of ABA (10 M) induced marked modification of embryo metabolism. Indeed, ABA completely prevented precocious embryo germination and inhibited lipid reserve catabolism. Moreover, the formation of small vacuoles and proliferation of rough endoplasmic reticulum in their vicinity suggested the onset of storage protein accumulation. After 15 days in the presence of ABA, the embryos contained abundant lipid and protein bodies. Nevertheless, these somatic embryos were not exactly the same as their mature zygotic counterparts since differences were found in chloroplasts, amyloplasts, and nuclear structures. These observations suggest that additional factors might be required to obtain fully mature somatic embryos.Abbreviations ABA abscisic acid - ABM ABA medium - BM basal medium - LB lipid bodies - MS Murashige and Skoog (1962) - PB protein bodies - RER rough endoplasmic reticulum     

Abscisic acid and osmotic induction of synchronous somatic embryo development of sweet potato

Summary Somatic embryos of sweet potato have potential as synthetic seeds. The effects of abscisic acid (ABA) (0,0,0.1, 1.0, 10.0 and 50.0 μM) were examined to improve synchrony and proliferation of somatic embryos. Transferring embryos compared to those cultures transferred at day 0. The development of embryos in suspension culture supplemented with ABA was poor. However, when calli proliferation cultures were in gelled medium and pulsed with 0.1 μM ABA for 14 d, the number of somatic embryos increased. Proembryonic masses cultured in mannitol-containing medium (Y=−1.5 MPa) increased embryo development and synchrony of embryo development. Thus, in this work ABA and mannitol have been shown to improve both the total number and the synchrony of sweet potato somatic embryos.     

Effects of ABA on secondary embryogenesis from somatic embryos induced from inflorescence culture ofAralia cordata Thunb

In order to investigate the effect of ABA on secondary embryogenesis from somatic embryos inAralia cordata Thunb., embryogenic callus and somatic embryos were induced from inflorescence on solid MS basal medium supplemented with 1.5 mg/L 2,4-D after eight weeks without subculture. For mass production of somatic embryos, embryogenic cell clumps were maintained in liquid MS medium supplemented with 1.0 mg/L 2,4-D, and then transferred to 2, 4-D-free medium. When developing embryos at various stages were cultured separately in liquid medium with ABA (0 to 2.0 mg/L) for three weeks, and then cultured in ABA-free liquid medium for two weeks, torpedo-shaped embryos exhibited secondary embryogenesis of 65.9% in only 0.2 mg/L ABA pretreatment. Cotyledonary embryos in cultures by 0.2, 0.5 and 1.0 mg/L ABA pretreatment also exhibited secondary embryogenesis (73%, 9.4% and 6.0%, respectively). However, globular and heart-shaped somatic embryos treated with ABA did not form secondary embryos on their hypocotyl surfaces. When cotyledonary embryos were cultured in ABA-free medium or 0.2 mg/L ABA treated medium for three weeks, and then in ABA-free liquid medium for 6 weeks, the germination frequency was lower in medium with 0.2 mg/L ABA (45.9%) than in hormone-free medium (56.8%). This result seems to be related to the high frequency of secondary embryogenesis. It is suggested that secondary embryogenesis by ABA application depends upon the stage of embryo cultured and the ABA concentration.     

Plant regeneration via direct somatic embryogenesis in Panax japonicus

Panax japonicus is one of the important medicinal plants. Here, we established the protocol for plant regeneration of P. japonicus via direct somatic embryogenesis. Somatic embryos were directly obtained from the segments of zygotic embryos on MS medium with 4.4 μM 2,4-D. Thereafter, somatic embryos were produced by repetitive secondary somatic embryogenesis. The secondary somatic embryo formation was enhanced by plasmolyzing pretreatment (1.0 M mannitol for 10 h). Frequency of secondary somatic embryo formation from cotyledon segments was lowered by plasmolyzing pretreatment, but the number of somatic embryos per explants was greatly increased. Plasmolyzing pretreatment resulted in retardation of embryo growth and required subculture to fresh medium for further growth of embryos into cotyledonary stage. Without plasmolyzing pretreatment, cotyledonary embryos were obtained after 8 weeks of culture. All the cotyledonary somatic embryos germinated by 5 μM GA₃ treatment, but only 15.3% were germinated on hormone-free medium. After 2 months of culture on 1/2 strength WPM medium, plantlets produced flowers spontaneously. In the anthers of in vitro flowers, microsporogenesis occurred normally with low number of pollen grains.     

Dormancy in somatic embryos and seeds ofVitis: changes in endogenous abscisic acid during embryogeny and germination

Abscisic acid (ABA) in extracts of somatic embryos and seeds of Gloryvine (Vitis vinifera L.xV. rupestris Scheele) was measured by gas chromatography-mass spectrometry-selected ion monitoring using deuterated ABA, (±)-[C-3Me-²H₃]ABA, ([²H₃]ABA) as internal standard. The ABA content increased rapidly during embryogeny (0.035 ng/embryo at the globular stage to 0.22 ng/embryo at the mature stage). The level of ABA in the tissues of somatic embryos, expressed in ng/mg dry weight, decreased from the globular stage (0.76 ng/mg) to the mature stage (0.25 ng/mg). Chilling (4° C) induced normal germination of seeds and mature somatic embryos and precocious germination of globular, heart-shaped and torpedoshaped somatic embryos. In all cases chilling led to a marked reduction in endogenous ABA. Exogenous (±)-ABA inhibited the germination of chilled somatic embryos.Abbreviations ABA abscisic acid - [²H₃]ABA (±)-[C-3Me-²H₃]-abscisic acid - BHT 2,6-di-t-butyl-4-methylphenol - GC-MS gas chromatography-mass spectrometry - Me-ABA and Me-[²H₃]ABA methyl esters of ABA and [²H₃]ABA, respectively - SIM selected ion monitoring     

Modulation of germination of embryos isolated from dormant and nondormant barley grains by manipulation of endogenous abscisic acid

Dormant and non-dormant barley (Hordeum distichum L.) grains with identical genetic backgrounds were obtained by maturing grains under different climate conditions. When isolated embryos from dormant grains were incubated in a well containing a fixed volume of water (300 l), the germination rate and percentage were dependent on the embryo number per well. A higher embryo number per well was correlated with a lower germination rate and percentage. However, this was not the case for the embryos isolated from nondormant grains. During germination, the endogenous cis-abscisic acid (ABA) in isolated embryos from both dormant and nondormant grains was analyzed. The inhibitory effect on germination of a higher number per well of isolated dormant embryos was due to diffusion of endogenous ABA out of the embryos and accumulation of ABA in the incubation medium. Moreover, there was de-novo synthesis of ABA in embryos isolated from dormant grains during incubation but not in embryos isolated from nondormant grains. The inhibitory effect of ABA on germination of embryos isolated from dormant grains could be mimicked by addition of ABA or the medium in which dormant embryos had been placed. Embryos isolated from nondormant grains were insensitive to addition of ABA and medium from dormant embryos. Our results demonstrate that diffusion of endogenous ABA, de-novo ABA synthesis and ABA sensitivity play a role in the control of germination. It is proposed that dormancy-breaking treatments act via changes to these processes.Abbreviations ABA cis-abscisic acid - E/W embryo(s) per well Prof. K.R. Libbenga (Institute of Molecular Plant Sciences, Leiden University) is thanked for fruitful discussions. B.V.D. was partly supported by E.E.C. BIOTECH program PL 920175.     

Scale-up of somatic embryogenesis in alfalfa (Medicago sativa L.) I subculture and indirect secondary somatic embryogenisis

Three methods of increasing the productivity of somatic embryogenesis in Medicago sativa L. were investigated. In the basic procedure, somatic embryos were initiated from young petioles and carried through several phases: callus formation, suspension culture, selection of the embryogenic fraction by sieving, development, maturation, desiccation and storage. The suspensions were normally separated into three fractions by sieving. Fraction I (<200 m) containing nonembryogenic cells or cell clusters was discarded. Fraction II (200–500 m) consisting of embryogenic cell clusters was collected for embryo development and maturation. Fraction III (over 500 M) containing the mixture of petiole residues with large pieces of calli and globular somatic embryos was usually discarded. Several methods to scale-up the suspension phase were unsuccessful. Direct subculture of the entire suspension by the addition of fresh liquid medium resulted in the loss of embryogenic capacity by the third subculture. Subculture of fraction II decreased embryogenic cell mass, and hence reduced total productivity. The recycling of fraction III back to fresh B₅g liquid medium resulted in high productivity in the first culture but further subculture of this fraction resulted in a rapid decline in the embryogenic capacity.As an alternative, somatic embryos from the first tissue culture cycle were also used as explants for the initiation of secondary embryogenic callus. The embryogenic capacity of these somatic embryo explants declined rapidly as they matured. More than 100 secondary somatic embryos could be induced from embryo explants removed from development medium at 10 days after sieving the suspension, but only 40 somatic embryos were produced from each mature somatic embryo explant, and 13 from desiccated embryos. The secondary somatic embryos were comparable to the primary embryos in quality according to germination tests. The implications of the results to the efficiency of somatic embryo production of Medicago are discussed.Abbreviations ABA abscisic acid - 2,4-d 2,4-dichlorophenoxyacetic acid - DAS days after sieving - PPF photosynthetic photon flux density - SE somatic embryo     

Effect of abscisic acid and stratification on somatic embryo maturation and germination of holm oak (<Emphasis Type="Italic">Quercus ilex</Emphasis> L.)

Summary The effect of abscisic acid (ABA) was evaluated during the maturation and germination of holm oak (Quercus ilex L.) somatic embryos. The addition of ABA to the culture medium significantly reduced unwanted recurrent embryogenesis in mature somatic embryos without affecting the germination of embryos subjected to stratification at 4°C. Stratification at 4°C for 2 mo. was the most efficient for stimulating somatic embryo germination of holm oak. The addition of 90 and 450 mM sucrose also improved germination, while higher sucrose concentrations were inhibitory.     

Somatic Embryogenesis in Abies Alba × Abies Alba and Abies Alba × Abies Nordmanniana Hybrids

Maturation and germination of somatic embryos of hybrids A. alba × A. alba and A. alba × A. nordmanniana were followed by protein analysis of single embryogenic -suspensor masses (ESM) and analysis of storage protein accumulation during somatic embryo development. Very important step was one week pre-cultivation of ESM on medium with polyethylene glycol (PEG) and abscisic acid (ABA). Low osmotic potential of maturation medium and addition of ABA supported development of somatic embryo. Also partial drying of somatic embryo during following three weeks was needed for its normal development. In spite of morphologically fully developed, the somatic embryos were not physiologically ready for germination at least in terms of storage protein accumulation.     

Seed development and vivipary in Zea mays L.

Seed development was investigated in kernels of developing wild-type and viviparous (vp-1) Zea mays L. Embryos and endosperm of wild-type kernels began to dehydrate at approx. 35 d after pollination (DAP); viviparous embryos did not desiccate but accumulated fresh weight via coleoptile growth in the caryopses. Concentrations of endogenous abscisic acid (ABA) in the embryo were relatively high early in development, being approx. 150 ng·g^-1 fresh weight at 20 DAP. The ABA content declined thereafter, falling to approx. 50 ng·g^-1 at 30 DAP. Endosperm ABA content was always low, being less than 20 ng·g^-1. There were no differences between wild-type and vp-1 tissues. Immature kernels did not germinate when removed from the ear until late in development. The ability to germinate was correlated with decreasing moisture content in the endosperm at the time of removal; premature drying of immature kernels resulted in greatly increased germination following imbibition. Excised embryos germinated precociously when removed from the endosperm as early as 25 DAP. Such germination could be prevented by treatment with 10^-5 M ABA or by lowering the solute potential (_s) of the medium with 0.3 M mannitol. Treatment of excised embryos with ABA led to internal ABA concentrations comparable to those in embryos in which germination was inhibited in situ. Mannitol treatment did not have this effect, although water-deficit stress of excised embryos resulted in substantial ABA production. Germinated vp-1 embryos were less sensitive to growth inhibition by ABA or mannitol than germinating wild-type embryos. The vp-1 seedlings were not wilty and their transpiration rates were reduced in response to ABA or water shortage.Abbreviations and symbols ABA abscisic acid - DAP days after pollination - FW fresh weight - vp-1 viviparous genotype - _s solute potential     

Effect of exogenous ABA on somatic embryo maturation and germination in Persian walnut (<Emphasis Type="Italic">Juglans regia</Emphasis> L.)

Low efficiency of embryo maturation, germination and conversion to plantlets is a major problem in many species including Persian walnut. We studied the effects of abscisic acid (ABA) and sucrose, on the maturation and germination of Persian walnut (Juglans regia) somatic embryos. Individual globular somatic embryos were grown on a maturation medium supplemented with different combinations of ABA and sucrose for ca. 1 month, until shoot meristems and radicles had developed. White and opaque embryos in late cotyledonary stage were subjected to desiccation after the culture period on maturation media. The number of germinated somatic embryos was influenced by the concentrations of ABA in the maturation medium. The best treatment for germination, in which both shoot and root were developed contained 2 mg l⁻¹ ABA and resulted in 41% conversion of embryos into plantlets. Regeneration was reduced at higher levels of ABA. While ABA always reduced the rate of secondary embryogenesis, treatments containing 4.0% sucrose significantly increased the number of secondary embryos. On the other hand, sucrose had little influence on maturation. Normal and abnormal embryos were verified anatomically.     

Onset of desiccation tolerance during development of the barley embryo

We have investigated events which take place in the developing barley (Hordeum vulgare L.) embryo during its acquisition of desiccation tolerance. Excised embryos are capable of precocious germination as early as 8 d after pollination (DAP). At this age, however, they are not capable of resisting a desiccation treatment which induces a loss of 96–98% of their initial water content. At 16 DAP the embryos germinate despite the drastic drying treatment. The pattern of in-vivo and in-vitro proteins synthesized by the developing embryos from 12 DAP (desiccation-intolerant) and 16 DAP (desiccation-tolerant) were compared. A set of 25–30 proteins was identified which is denovo synthesized or enhanced during the developmental period leading to desiccation tolerance. Abscisic acid (ABA; 100 M) applied in vitro for 5 d to 12-DAP embryos induces desiccation tolerance and represses a subset of polypeptides preferentially associated with 16-DAP embryos. During in vitro culture of barley embryos ABA stimulates the appearance of a set of proteins and prevents the precocious germination allowing embryogenesis to continue in vitro. It also suppresses a set of germination-related proteins which appear 4 h after the incubation of the dissected embryo on a germination medium without ABA. Almost all mRNAs remain functional for translation when isolated embryos are dried at the desiccation-intolerant and tolerant stages of embryo development.Abbreviations ABA abscisic acid - DAP days after pollination - GM germination medium - poly(A)RNA polyadenylated RNA - SDS sodium dodecyl sulfate     

Proliferation and maintenance of embryogenic capacity in elm embryogenic cultures

Summary This paper investigates maintenance and proliferation of somatic embryogenesis systems for Ulmus minor and U. glabra. Proliferation occurred with subculture of embryogenic calluses. The calluses were mainly formed by friable nodules composed of meristematic cells organized into proembryogenic cell masses (PEMs) and thin-walled vacuolated parenchymatic cells. Cotyledonary embryos, with procambial strands and differentiation of their vascular tissues as well as visible root meristems, were identifiable after 18d of culture on a proliferation medium with 0.44 μM benzyladenine (BA). The shoot meristem was only occasionally well developed. Somatic embryo multiplication from elm embryogenic calluses is a clearly asynchronic system, and PEMs as well as embryos at all stages of development are observed simultaneously at the end of subculture period. Factors affecting the proliferation of elm embryogenic callus, such as culture medium, carbon source and genotype, were studied. Basal medium (MS) or medium supplemented with 0.44 μM BA produced the highest number of somatic embryos. Somatic embryo production was higher with sucrose or glucose than with maltose, and significant differences were also found among the four embryogenic lines tested. The use of liquid medium with filter paper support is an essential step for the survival of isolated somatic embryos during the germination stage. The addition of 0.22 μM BA′ to liquid MS medium was the best treatment for germination and plantlet conversion of elm somatic embryos.     

Some features of somatic embryo maturation of algerian fir

Summary Maturation of Abies numidica De Lann. somatic embryos was tested on media with abseisic acid and various maltose and polyethylene glycol-4000 concentrations. The effect of basal medium and subculture period on maturation was also examined. The maturation of somatic embryos was promoted by polyethylene glycol-4000, at 7.5–10%. Three to 6% maltose significantly enhanced the yield of mature embryos. The most effective somatic embryo maturation occurred when embryogenictissue was transferred to maturation medium after 14–21 d cultivation on proliferation medium. The ability for A. numidica cultures to form cotyledonary somatic embryos was assessed over a period of 3 yr. Plantlets germinated on half-strength SH (Schenk and Hildebrandt) medium with charcoal and survived transfer to the soil.